A major pathway of eukaryotic messenger RNA (mRNA) turnover begins with deadenylation, followed by decapping and 5′ to 3′ exonucleolytic decay. We provide evidence that mRNA decapping and 5′ to 3′ degradation occur in discrete cytoplasmic foci in yeast, which we call processing bodies (P bodies). First, proteins that activate or catalyze decapping are concentrated in P bodies. Second, inhibiting mRNA turnover before decapping leads to loss of P bodies; however, inhibiting turnover at, or after, decapping, increases the abundance and size of P bodies. Finally, mRNA degradation intermediates are localized to P bodies. These results define the flux of mRNAs between polysomes and P bodies as a critical aspect of cytoplasmic mRNA metabolism and a possible site for regulation of mRNA degradation.Messenger RNA turnover is an important step in regulating gene expression. In yeast, the major pathway of mRNA decay is initiated by shortening of the 3′ poly(adenosine) [poly(A)] tail, followed by removal of the cap by the decapping enzyme Dcp1p/Dcp2p, which in turn allows 5′ to 3′ exonucleolytic decay (1-8). Decapping is a key step in this pathway, because it permits the destruction of the mRNA and is a site of numerous control inputs (9).Several observations suggest that decapping occurs when the mRNA undergoes a transition from a translationally competent messenger ribonucleoprotein (mRNP) to an mRNP state destined for decay. For example, the translation initiation factor eIF4E, which binds the cap structure, is an inhibitor of decapping both in vitro and in vivo (10,11). Moreover, deadenylated mRNAs that interact with a complex of Lsm1-7 proteins, which activates decapping, are no longer bound by eIF4E or eIF4G (12).The hypothesis that mRNAs enter a non-translating state after deadenylation and before decapping is analogous to the storage of mRNA in numerous biological contexts where deadenylated mRNAs are translationally repressed before their later activation. Consistent with a mechanistic similarity between decapping and mRNA storage, Dhh1p, which is an activator of decapping in yeast (13), has homologs that are required for the translational repression and storage of maternal mRNAs in Xenopus, Drosophila, and Caenorhabditis (14-16). Such stored mRNAs are often localized in discrete cytoplasmic granules, which represent accumulations of translationally repressed mRNAs (15). This analogy suggested the possibility that Dhh1p

Recent results indicate that many untranslating mRNAs in somatic eukaryotic cells assemble into related mRNPs that accumulate in specific cytoplasmic foci referred to as P bodies. Transcripts associated with P body components can either be degraded or return to translation. Moreover, P bodies are also biochemically and functionally related to some maternal and neuronal mRNA granules. This suggests an emerging model of cytoplasmic mRNA function in which the rates of translation and degradation of mRNAs are influenced by a dynamic equilibrium between polysomes and the mRNPs seen in P bodies. Moreover, some mRNA-specific regulatory factors, including miRNAs and RISC, appear to repress translation and promote decay by recruiting P body components to individual mRNAs.

Recent experiments have defined cytoplasmic foci, referred to as processing bodies (P-bodies), wherein mRNA decay factors are concentrated and where mRNA decay can occur. However, the physical nature of P-bodies, their relationship to translation, and possible roles of P-bodies in cellular responses remain unclear. We describe four properties of yeast P-bodies that indicate that P-bodies are dynamic structures that contain nontranslating mRNAs and function during cellular responses to stress. First, in vivo and in vitro analysis indicates that P-bodies are dependent on RNA for their formation. Second, the number and size of P-bodies vary in response to glucose deprivation, osmotic stress, exposure to ultraviolet light, and the stage of cell growth. Third, P-bodies vary with the status of the cellular translation machinery. Inhibition of translation initiation by mutations, or cellular stress, results in increased P-bodies. In contrast, inhibition of translation elongation, thereby trapping the mRNA in polysomes, leads to dissociation of P-bodies. Fourth, multiple translation factors and ribosomal proteins are lacking from P-bodies. These results suggest additional biological roles of P-bodies in addition to being sites of mRNA degradation.